Use of tethering for axial confinement in optical tweezers

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Use of tethering for axial confinement in optical
tweezers

• Mark Cronin-Golomb
• Biomedical Engineering
• Tufts University

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Outline

• Motivation
• Design of l DNA tether
• Videos of untethered and tethered particles
• Confocal detection measurement system
• Demonstration of force measurement
• Future directions

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Tethers and tweezers

• Microspheres tethered to each other (Chu)
• Backscattering from tethered bead as probe of DNA
  flexibility (Libchaber APL 73, 291 (1998))
• Twisting polymers by applying torque to trapped
  particle (Bustamante Nature 424, 338 (2003),
  Ormos)
• Study of macromolecular motion (Gelles)

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Use of low numerical aperture trapping lenses

• Trapping particles against glass slide
• Trapping against counterflow
• Trapping against gravity

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Axial trapping is harder to achieve than
transverse trapping

• Generalized Lorenz-Mie theory to find radiation
  pressure cross section Cpr(z) and radiation
  pressure force F in terms of standard Mie
  scattering coefficients

K.F. Ren, G. Gréhan, and G. Gouesbet, Appl. Opt.
35, 2702 (1996)
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Axial force with 1.25NA beam
1mm diameter polystyrene bead, 13mW 820nm
wavelength trap
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Axial force for 0.65NA beam
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Beads in 0.65NA trap without tether
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Comparison of original and tethered configurations
l DNA 48k base pairs 31.5x106 Dalton
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Experiment Details
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End labeling l DNA for attachment to streptavidin
and anti-digoxigenin
1. dNTPs dCTP biotin-dUTP Klenow
2. dCTP digoxigenin-dUTP
Zimmermann and Cox, Nucleic Acids Research 22,
492 (1994)
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Tether construct
Modified from Meiners and Quake Phys. Rev. Lett.
84, 5014 (2000)
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Frame sequence from tethered bead video
10 mm
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Tethered beads in 0.65NA trap
Tracking Software implemented in IDL by Crocker
and Weeks http//www.physics.emory.edu/weeks/idl/

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Experiment Details measurements
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• As the tweezer beam is moved back and forth, the
  probe bead lags behind.
• The bead is bright when the tweezer beam
  illuminates it.
• The confocal signal is highest when the tweezer
  beam is centered on the probe bead.

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At large oscillation amplitudes the potential
well splits
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Theoretical Background
x trap position g viscous drag k tweezer
spring constant a amplitude of trap
oscillation w frequency of trap oscillation
L(t) Brownian forcing function
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Viscosity Image

• Viscosity distribution around A. pullulans imaged
  by raster scanning an optically trapped probe
  bead.
• This blastospore has a halo of the polysaccharide
  pullulan around it. Note the viscosity gradient.

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Force Off
Force On
Probe Bead
Probe Bead
r
a
a
Oscillating Laser Trap
Oscillating Laser Trap
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Force Measurement

• We can use confocal tweezers to measure forces
  applied to probe beads.
• Flow measurement is one example of force
  measurement

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Force measurement

• An optically trapped microsphere is used as a
  probe for two-dimensional force imaging using
  scanning optics.
• A fluid viscosity map may be obtained
  simultaneously.
• Calibration is based on a single length
  measurement only the oscillation amplitude a of
  the trap.

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Transverse force on tethered bead
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Further applications

• Fiber based sensor

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Applications

• Photonic force microscope with retained probe
  bead
• Measurement of changes in tether properties with
  environment, e.g. with enzymes, buffer properties
  etc.

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Array of tethered beads for actin network network
generation and analysis
Actin
From Christian Schmitz talk
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Conclusions

• Probe beads can be tethered to substrates to
  eliminate need for axial trapping, enabling use
  of low NA objectives.
• Measurements of viscosity and force can be made
  with tethered beads via confocal detection system
• References to confocal detection method
• Nemet, Shabtai, Cronin-Golomb, Opt. Lett. 27, 264
  (2002)
• Nemet, Cronin-Golomb, Opt. Lett. 27, 1357, (2002)
• Nemet, Cronin-Golomb, Appl. Opt. 42, 1820 (2003)

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Acknowledgements

• Boaz Nemet
• Joe Platko
• Support of Tufts University Bioengineering Center